Project Summary/Abstract It has been more than 50 years since Eugene Kennedy described/identified phosphatidic acid phosphatase (PAP) activity. This enzymatic activity unified what is now known as the Kennedy pathway of phosphatidylcholine synthesis. The genes encoding PAP, termed Lipin 1-3, sit at an important branch point in the Kennedy pathway of glycerolipid synthesis. Unlike the remainder of the enzymes in the triacylglycerol synthesis pathway, the lipins are cytosolic proteins that translocate from the cytosol to their site of action at internal membranes. Loss of function alleles of lipin family members results in profound metabolic and inflammatory disturbances in both mice and humans, and lipin polymorphisms are linked to a number of metabolic conditions. But while genetics has demonstrated the importance of these enzymes in health and disease, precisely how they are regulated is still poorly defined. Elucidating the mechanisms and pathways controlling lipin family activity will provide insight into glycerolipid biosynthesis and the pathophysiological consequences when it is disrupted, and will open new avenues for pharmacological interventions for these diseases. This proposal undertakes a rigorous and comprehensive examination of lipin 1 regulation with the goal of understanding how its PAP activity is biochemically controlled. We will characterize the effects of two kinases on lipin 1 PAP activity and stability and identify how these regulatory events occur. In addition, we will determine how dysregulation of lipin PAP activity impacts cellular function, particularly neutral and phospholipid synthesis. Given the striking phenotypes displayed by genetic alterations in the lipins, accomplishing these aims will yield important insights into lipid biosynthetic pathways as well as clinically relevant pathologies.